Non-linear response of Norway spruce to climate variation along elevational and age gradients in the Carpathians.

Climate change, namely increased warming coupled with a rise in extreme events (e.g., droughts, storms, heatwaves), is negatively affecting forest ecosystems worldwide. In these ecosystems, growth dynamics and biomass accumulation are driven mainly by environmental constraints, inter-tree competition, and disturbance regimes. Usually, climate-growth relationships are assessed by linear correlation due to the simplicity and straightforwardness of modeling. However, applying this method may bias results, since the ecological and physiological responses of trees to environmental factors are non-linear, and usually bell-shaped. In the Eastern Carpathian, Norway spruce is at the southeasternmost edge of its natural occurrence; this region is thus potentially vulnerable to climate change. A non-linear assessment of climate-growth relationships using machine-learning techniques for Norway spruce in this area had not been conducted prior to this study. To address this knowledge gap, we analyzed a large tree-ring network from 158 stands, with over 3000 trees of varying age distributed along an elevational gradient. Our results showed that non-linearity in the growth-climate response of spruce was season-specific: temperatures from the previous autumn and current growing season, along with water availability during winter, induced a bell-shaped response. Moreover, we found that at low elevations, spruce growth was mainly limited by water availability in the growing season, while winter temperatures are likely to have had a slight influence along the entire elevational gradient. Furthermore, at elevations lower than 1400 m, spruce trees were also found to be sensitive to previous autumn water availability. Overall, our results shed new light on the response of Norway spruce to climate in the Carpathians, which may aid in management decisions.

Detecting changes in industrial pollution by analyzing heavy metal concentrations in tree-ring wood from Romanian conifer forests.

The impact of air pollution on forests, especially in urban areas, has been increasingly discussed recently. Many pollutants, including heavy metals, are released into the atmosphere from various sources, such as mining, non-ferrous metal processing plants, and fossil fuel combustion. These pollutants can adversely affect not only tree growth but also other species, including humans. This study compared the concentrations of several elements in tree-ring wood from two conifer species (Silver fir, Abies alba; Norway spruce, Picea abies) growing in polluted and unpolluted areas. Two regions in northern Romania (Bicaz and Tarnița) that were subjected to historical pollution changes were selected. Two chemical analyses were used: inductively coupled plasma mass spectrometry (ICP-MS) and X-ray fluorescence spectrometry (XRF). The silver fir trees from the intensively polluted area in the Tarnița region were negatively impacted by industrial pollution: the Mn concentrations were, on average, three times higher in polluted areas than in unpolluted areas (ca. 30 vs. 10 mg kg-1). This finding was consistent for both ICP-MS and XRF analyses. However, in Norway spruce, this difference was found only in the XRF data, which detected Mn concentrations seven times higher in trees from polluted areas than those from unpolluted areas (ca. 700 vs. 100 mg kg-1). In the Tarnița region, Norway spruce accumulated more heavy metals than silver fir, but the most pronounced differences between polluted and unpolluted areas were found in silver fir. The two analytical methods are commonly used to determine metal concentrations in wood, and they complement each other, with ICP-MS having a low detection limit for some elements and XRF having higher detection limits and better accuracy. Each method has its advantages and disadvantages, and the optimal method depends on many factors, such as the type of heavy metal analyzed, its concentration in wood, sample type, cost, analysis time, and sample preparation.

Strategic roadmap to assess forest vulnerability under air pollution and climate change.

Although it is an integral part of global change, most of the research addressing the effects of climate change on forests have overlooked the role of environmental pollution. Similarly, most studies investigating the effects of air pollutants on forests have generally neglected the impacts of climate change. We review the current knowledge on combined air pollution and climate change effects on global forest ecosystems and identify several key research priorities as a roadmap for the future. Specifically, we recommend (1) the establishment of much denser array of monitoring sites, particularly in the South Hemisphere; (2) further integration of ground and satellite monitoring; (3) generation of flux-based standards and critical levels taking into account the sensitivity of dominant forest tree species; (4) long-term monitoring of N, S, P cycles and base cations deposition together at global scale; (5) intensification of experimental studies, addressing the combined effects of different abiotic factors on forests by assuring a better representation of taxonomic and functional diversity across the ~73,000 tree species on Earth; (6) more experimental focus on phenomics and genomics; (7) improved knowledge on key processes regulating the dynamics of radionuclides in forest systems; and (8) development of models integrating air pollution and climate change data from long-term monitoring programs.

Allocation of Nutrients and Leaf Turnover Rate in Poplar under Ambient and Enriched Ozone Exposure and Soil Nutrient Manipulation.

An excess of ozone (O3) is currently stressing plant ecosystems and may negatively affect the nutrient use of plants. Plants may modify leaf turnover rates and nutrient allocation at the organ level to counteract O3 damage. We investigated leaf turnover rate and allocation of primary (C, N, P, K) and secondary macronutrients (Ca, S, Mg) under various O3 treatments (ambient concentration, AA, with a daily hourly average of 35 ppb; 1.5 × AA; 2.0 × AA) and fertilization levels (N: 0 and 80 kg N ha-1 y-1; P: 0 and 80 kg N ha-1 y-1) in an O3-sensitive poplar clone (Oxford: Populus maximowiczii Henry × P. berolinensis Dippel) in a Free-Air Controlled Exposure (FACE) experiment. The results indicated that both fertilization and O3 had a significant impact on the nutrient content. Specifically, fertilization and O3 increased foliar C and N contents (+5.8% and +34.2%, respectively) and root Ca and Mg contents (+46.3% and +70.2%, respectively). Plants are known to increase the content of certain elements to mitigate the damage caused by high levels of O3. The leaf turnover rate was accelerated as a result of increased O3 exposure, indicating that O3 plays a main role in influencing this physiological parameter. A PCA result showed that O3 fumigation affected the overall allocation of primary and secondary elements depending on the organ (leaves, stems, roots). As a conclusion, such different patterns of element allocation in plant leaves in response to elevated O3 levels can have significant ecological implications.

Monitoring of ozone in selected forest ecosystems in Southern Carpathian and Romanian Intensive Monitoring Network (level II).

In the Romanian forest ecosystems, the first measurements of ambient ozone (O(3)) concentrations started in 1997 in 6 of 26 locations established in a trans-Carpathian Network. Furthermore, three additional ozone and other phytotoxic pollutant (NO(x), SO(2) and NH(3)) monitoring networks were installed in 2000 in Retezat (11 locations) and during 2006-2009 in Bucegi-Piatra Craiului (22 locations) LTER Sites. Since 2007, in four Intensive Forest Monitoring plots (level II), measurements of ozone concentrations were developed. Measurements were made using the Ogawa® passive sampler system during the growing season (April to October). In the Bucegi LTER Site, the seasonal means of 42.5-47.2 ppb in 2006 and 2008 were higher than those determined in the Carpathian Network in the 1997-1999 period (39.0-42.0 ppb), while the 2009 mean of 40.0 ppb was in the range of these values. The O(3) levels were slightly higher than those measured in Retezat LTER Site. In the Intensive Forest Monitoring Network (level II), no significant differences in ozone concentrations between individual core plots were noticed. The seasonal means for each plot range between 36.8 and 49.8 ppb in 2008. An influence of ozone concentrations on crown condition and tree volume growth was not determined.

Status of the Southern Carpathian forests in the long-term ecological research network.

Air pollution, bulk precipitation, throughfall, soil condition, foliar nutrients, as well as forest health and growth were studied in 2006-2009 in a long-term ecological research (LTER) network in the Bucegi Mountains, Romania. Ozone (O(3)) was high indicating a potential for phytotoxicity. Ammonia (NH(3)) concentrations rose to levels that could contribute to deposition of nutritional nitrogen (N) and could affect biodiversity changes. Higher that 50% contribution of acidic rain (pH < 5.5) contributed to increased acidity of forest soils. Foliar N concentrations for Norway spruce (Picea abies), Silver fir (Abies alba), Scots pine (Pinus sylvestris), and European beech (Fagus sylvatica) were normal, phosphorus (P) was high, while those of potassium (K), magnesium (Mg), and especially of manganese (Mn) were significantly below the typical European or Carpathian region levels. The observed nutritional imbalance could have negative effects on forest trees. Health of forests was moderately affected, with damaged trees (crown defoliation >25%) higher than 30%. The observed crown damage was accompanied by the annual volume losses for the entire research forest area up to 25.4%. High diversity and evenness specific to the stand type's structures and local climate conditions were observed within the herbaceous layer, indicating that biodiversity of the vascular plant communities was not compromised.

Visible Foliar Injury and Ecophysiological Responses to Ozone and Drought in Oak Seedlings.

To verify the responses of visible foliar injury (VFI), we exposed seedlings of three oak species for 4.5 months in an open air facility, using differing ozone (O3) and drought treatments: O3 (three levels from ambient to ×1.4 ambient), and drought (three levels of irrigation from 40% to 100% field capacity). We related the accumulated phytotoxic O3 dose (POD1) and cumulative drought index (CDI) to the O3 and drought VFI and assessed growth increment (height, diameter, leaf number), biomass (of all organs), and physiological parameters: net photosynthesis per plant (Pn), photosynthetic nitrogen (PNUE) and phosphorus use efficiency (PPUE)). The results indicated that an increase in POD1 promoted O3 VFI in Quercus robur and Quercus pubescens, while Quercus ilex was asymptomatic. The POD1-based critical level at the onset of O3 VFI was lower for Q. robur than for Q. pubescens (12.2 vs. 15.6 mmol m-2 POD1). Interestingly, drought reduced O3 VFI in Q. robur but increased it in Q. pubescens. Both O3 and drought were detrimental to the plant biomass. However, Q. robur and Q. pubescens invested more in shoots than in roots, while Q. ilex invested more in roots, which might be related to a hormetic mechanism. Pn, PNUE and PPUE decreased in all species under drought, and only in the sensitive Q. robur (PPUE) and Q. pubescens (PNUE) under O3. This study confirms that POD1 is a good indicator to explain the development of O3 VFI and helps a differential diagnosis of co-occurring drought and O3 VFI in oak forests.

Do Different Tree-Ring Proxies Contain Different Temperature Signals? A Case Study of Norway Spruce (Picea abies (L.) Karst) in the Eastern Carpathians.

One of the most important proxy archives for past climate variation is tree rings. Tree-ring parameters offer valuable knowledge regarding how trees respond and adapt to environmental changes. Trees encode all environmental changes in different tree-ring parameters. In this study, we analyzed how air temperature is encoded in different Norway spruce tree-ring proxies along an altitude gradient in an intramountain valley of the Carpathians. The study area, in the Gheorgheni region, Romania (Eastern Carpathians), has a mountain climate with a frequent temperature inversion in winter. The climate−growth relationship was analyzed for two contrasting altitudes: low elevation, i.e., below 1000 m a.s.l., and high elevation, i.e., above 1500 m a.s.l. Two local weather stations, one in the valley and the other on the upper part of the mountains, provide daily temperatures (Joseni­750 m a.s.l. and Bucin­1282 m a.s.l.). The bootstrap Pearson correlation between cumulative daily temperature data and three tree-ring proxies (tree-ring width­TRW, basal area increment­BAI, and blue intensity­BI) was computed for each series. The results show that elevation modulates the climate response pattern in the case of BI, and remains relatively similar for TRW and BAI. The winter temperature's positive influence on spruce growth was observed in both TRW and BAI chronologies. Additionally, the BAI chronology highlights a positive relationship with summer temperature. The highest correlation coefficient (r = 0.551, p < 0.05, n = 41) was recorded between BI residual chronology from high elevation series and summer/autumn temperature from the upper-part weather station for a cumulative period of 59 days (the second half of August to the beginning of October). Our results show that, for this intramountain valley of the Eastern Carpathians, different tree-ring proxies capture different climate signals.

Species discrimination and individual tree detection for predicting main dendrometric characteristics in mixed temperate forests by use of airborne laser scanning and ultra-high-resolution imagery.

This study aims to investigate the combined use of two types of remote sensing data - ALS derived and digital aerial photogrammetry data (based on imagery collected by airborne UAV sensors) - along with intensive field measurements for extracting and predicting tree and stand parameters in even-aged mixed forests. The study is located in South West Romania and analyzes data collected from mixed-species plots. The main tree species within each plot are Norway spruce (Picea abies L. Karst.) and Beech (Fagus sylvatica L.). The ALS data were used to extract the digital terrain model (DTM), digital surface model (DSM) and normalized canopy height model (CHM). Object-Based Image Analysis (OBIA) classification was performed to automatically detect and separate the main tree species. A local filtering algorithm with a canopy-height based variable window size was applied to identify the position, height and crown diameter of the main tree species within each plot. The filter was separately applied for each of the plots and for the areas covered with Norway spruce and beech trees, respectively (i.e. as resulted from OBIA classification). The dbh was predicted based on ALS data by statistical Monte Carlo simulations and a linear regression model that relates field dbh for each tree species with their corresponding ALS-derived tree height and crown diameter. The overall RMSE for each of the tree species within all the plots was 5.8 cm for the Norway spruce trees, respectively 5.9 cm for the beech trees. The results indicate a higher individual tree detection rate and subsequently a more precise estimation of dendrometric parameters for Norway spruce compared to beech trees located in spruce-beech even-aged mixed stands. Further investigations are required, particularly in the case of choosing the best method for individual tree detection of beech trees located in temperate even-aged mixed stands.

Climate change effects on tree growth from Romanian forest monitoring Level II plots.

Forest health status is negatively influenced by climate change, air pollution and other disturbances. Extreme droughts reduce stand productivity, increase vulnerability to pests, and can even provoke mortality. Growth dynamics at tree and forest stand levels are considered the main indicators of stability and productivity in forest ecosystem structures. The main climate drivers for tree growth were identified using basal area increment (BAI) as a synthetic indicator. BAI chronologies were obtained from increment cores for 1960-2012 period. Six species were analysed in an attempt to identify their growth limiting factors. For the most important oak species in Romania, resilience components were computed in order to analyse their response to drought events. Moreover, growth dynamics were analysed for two species in mixed and monoculture forests. The results suggest that - in comparison to Picea abies and Fagus sylvatica, the sensitivity of Quercus spp. is much higher (0.3-0.47). Oakspecies situated in the most drought-affected areas are sensitive to rainfall values from the previous autumn, current spring, and early summer, with April monthly values having the most significant effect on BAI increment (r = 0.47*) The most sensitive species to drought is Q. cerris and Q. frainetto. Their BAI reduction during drought is >50% compared with the BAI values before the drought period. The recovery capacity of tree growth following drought events is lower for Q. robur and Q. petraea and higher for Q. cerris and Q. frainetto. The mixed forest stands have not showed a constant higher resistance to drought.

Ozone impairs the response of isoprene emission to foliar nitrogen and phosphorus in poplar.

Tropospheric ozone (O3) impairs physiological processes of plants while nitrogen (N) deposition may cause imbalances in soil N and other nutrients such as phosphorus (P) suggesting an increase of P demand for plants. However, the combined effect of O3, soil N and P on isoprene emission from leaves has never been tested. We therefore examined isoprene emission in leaves of Oxford poplar clone exposed to O3 (ambient, AA [35.0 nmol mol-1 as daily mean]; 1.5 × AA; 2.0 × AA), soil N (0 and 80 kg N ha-1) and soil P (0, 40 and 80 kg P ha-1) in July and September in a Free-Air Controlled Exposure (FACE) facility. We also investigated the response of isoprene emission to foliar N, P and abscisic acid (ABA) contents in September because the 2-C-methylerythritol-5-phosphate (MEP) pathway of isoprenoid biosynthesis produces ABA. We found that O3 increased isoprene emission in July, which was associated to increased dark respiration, suggesting an activation of metabolism against O3 stress as an initial response. However, O3 decreased isoprene emission in September which was associated to reduced net photosynthesis. In September, isoprene emission was positively correlated with leaf N content and negatively correlated with leaf P content in AA. However, no response of isoprene emission to foliar N and P was found in elevated O3, suggesting that the isoprene responses to foliar N and P depended on the O3 exposure levels. Isoprene emission rate in 1.5 × AA and 2.0 × AA increased with increasing leaf ABA content, indicating accelerated senescence of injured leaves to favor new leaf growth when high O3 and nutritional availability in the soil were combined. Even though foliar N and P usually act as a proxy for isoprene emission rate, the impact of recent abiotic factors such as O3 should be always considered for modeling isoprene emission under climate change.

Estimating forest stand structure attributes from terrestrial laser scans.

Forest stands are often parameterized by vegetation indices such as the Leaf Area Index (LAI). However, other indices (i.e. stand denseness, espacement, canopy density, canopy cover, foliage cover, crown porosity, gap fraction) may better characterize forest structure. Terrestrial and airborne active sensor data has been used to describe canopy structural diversity and provide accurate estimates of forest structure indices. This study uses Terrestrial Laser Scanner (TLS) to characterize forest structure through the above-mentioned indices. The relationship between all of them was studied to assess the extent to which they relate and their capability to properly describe forest stands. A strong correlation was visible between LAI and the canopy density index (r = 0.87 to 0.91 depending on the extraction methods) despite the underevaluated values of the first. Even though more precise LAI estimates were expected from using co-registered multiple scans, the LAI variability proved to be low and correlations with the remaining indices weakened when compared to a single scan approach. An exception was canopy cover, a structural index that disregards the three-dimensionality of the canopy, with which the LAI obtained from multiple scans maintained a strong correlation. This suggests that multiple scanning leads to an unweighted oversampling of the scene, overshadowing its advantages in removing tree occlusions. Weak correlations were visible between classic forest structural indices (basal area density index, espacement index, denseness index) and the rest of the descriptors. Despite this exception, most of the forest indices showed average to strong correlations in-between each other. Therefore, we conclude that a better description of forest stands structure may be achieved through unsegmented single scan point cloud processing in both 3D and 2D space, optical data from the incorporated digital camera being a plus, but not an essential requirement.

Testing three climate datasets for dendroclimatological studies of oaks in the South Carpathians.

Three gridded datasets containing interpolated daily and monthly precipitation and temperature values over the past five decades were tested against four tree-ring chronologies of oak (Quercus robur and Q. petraea). The objective of this research was to investigate the climate-growth relationship and whether the Pearson's product-moment correlation coefficients differ significantly if mean monthly precipitation and temperature data from the different climate databases, CRU, E-OBS and ROCADA are used. To this end, we selected two representative oak ecosystems in the South Carpathians, Romania, and analysed earlywood, latewood and tree-ring widths. Climate time series trends for the South Carpathians coldest, warmest days and wettest days were assessed with datasets from E-OBS and ROCADA, which differed in the density of their meteorological station network and their interpolation methods. The observed climatic parameters showed changes towards wetter conditions after the mid-1980s. For 1961-2013, E-OBS underestimated the mean daily temperature and daily precipitation compared with ROCADA. The results showed that higher extreme temperatures from January-March affected earlywood growth. In the investigated study region, latewood formation seemed to be affected by water availability mainly in May. Periods of drought associated with higher temperatures have limiting effects on tree growth, but these events are captured in different ways by each climate database analysed. Similarly, the results showed the discrepancy among datasets for earlywood and climate relationships. The results emphasize the importance of proper selection of climate data for assessing climate-tree growth relationships. For future dendroclimatological and dendroecological studies of oak in Romania, we recommend the ROCADA database, while E-OBS is recommended if an up-to-date climate dataset is needed.

Radial growth-based assessment of sites effects on pedunculate and greyish oak in southern Romania.

This study focuses on the climate growth drivers of Quercus robur L. (pedunculate oak) and Q. robur subsp. pedunculiflora K. Koch. (greyish oak), occurring in the biodiversity of three sites in southern Romania. We determined the degree of tolerance of the greyish oak, between the tardive and praecox varieties, to environmental stress, between 1951 and 2016. Total tree ring-width (RW), and earlywood (EW) and latewood (LW) measurements were subject of periodical and monthly climate-growth analysis. Our results revealed a moderate relationship between climate and tree-growth. A significant and positive relationship was observed between RW and previous growing season precipitation. Mean and minimum temperatures affected both positive and negative tree-rings during the growing season. We also observed that winter and spring represent key seasons for differentiating tardive from praecox varieties, affecting the intra-annual variability of ring-width, and EW and LW parameters. The correlation between the tree-ring measurements and daily climate data shows a clear offset of the starting growth between greyish oak varieties. A weak influence of stressors on tree-growth at the sites was observed through pointer year and resilience components analysis.

Forest vulnerability to extreme climatic events in Romanian Scots pine forests.

In the last years, large-scale mass forest withering and dieback have been reported for Scots pine (Pinus sylvestris) across eastern Europe, particularly in Romania. In these regions, the climate models forecast an increase in intensity and frequency of extreme climate events such as drought. Taking into account these aspects, the exact identification of the influences of drought on the loss of radial growth and vitality in Scots pine stands becomes mandatory. To achieve this aim, we developed the first country-wide Scots pine dendrochronological network in Romania consisting of 34 chronologies of basal area increment (BAI), and including 1401 individual tree-ring width series. Romanian Scots pine forests were severely impacted by the 2000 and 2012 droughts. The high temperatures and low precipitation from April to August were the main climatic causes of radial-growth reduction and large-scale withering in some areas. By mapping post-drought growth resilience, we identified locations where resilience was low and could identify foci of future forest dieback and high tree mortality. The projected appearance of similar prolonged and severe droughts in the future will lead to the damage or local extinction of some Scots pine forests in Romania, regardless of their age, composition or spatial location. The elaboration of adaptive forest management strategies to the impact of climate changes, specifically designed for the Scots pine stands, is not possible without knowing and understanding these aspects.

Cross-talk between physiological and biochemical adjustments by Punica granatum cv. Dente di cavallo mitigates the effects of salinity and ozone stress.

Plants are exposed to a broad range of environmental stresses, such as salinity and ozone (O3), and survive due to their ability to adjust their metabolism. The aim of this study was to evaluate the physiological and biochemical adjustments adopted by pomegranate (Punica granatum L. cv. Dente di cavallo) under realistic field conditions. One-year-old saplings were exposed to O3 [two levels denoted as ambient (AO) and elevated (EO) O3 concentrations] and salinity [S (salt, 50 mM NaCl)] for three consecutive months. No salt (NS) plants received distilled water. Despite the accumulation of Na+ and Cl- in the aboveground biomass, no evidence of visible injury due to salt (e.g. tip yellow-brown lesions) was found. The maintenance of leaf water status (i.e. unchanged values of electrolytic leakage and relative water content), the significant increase of abscisic acid, proline and starch content (+98, +65 and +59% compared to AO_NS) and stomatal closure (-24%) are suggested to act as adaptive mechanisms against salt stress in AO_S plants. By contrast, EO_NS plants were unable to protect cells against the negative impact of O3, as confirmed by the reduction of the CO2 assimilation rate (-21%), accumulation of reactive oxygen species (+10 and +225% of superoxide anion and hydrogen peroxide) and malondialdehyde by-product (about 2-fold higher than AO_NS). Plants tried to preserve themselves from further oxidative damage by adopting some biochemical adjustments [i.e. increase in proline content (+41%) and induction of catalase activity (8-fold higher than in AO_NS)]. The interaction of the two stressors induced responses considerably different to those observed when each stressor was applied independently. An analysis of the antioxidant pool revealed that the biochemical adjustments adopted by P. granatum under EO_S conditions (e.g. reduction of total ascorbate; increased activities of superoxide dismutase and catalase) were not sufficient to ameliorate the O3-induced oxidative stress.

Synthetic aperture radar sensitivity to forest changes: A simulations-based study for the Romanian forests.

Natural and anthropogenic disturbances pose a significant threat to forest condition. Continuous, reliable and accurate forest monitoring systems are needed to provide early warning of potential declines in forest condition. To address that need, state-of-the-art simulations models were used to evaluate the utility of C-, L- and P-band synthetic aperture radar (SAR) sensors within an integrated Earth-Observation monitoring system for beech, oak and coniferous forests in Romania. The electromagnetic simulations showed differentiated sensitivity to vegetation water content, leaf area index, and forest disturbance depending on SAR wavelength and forest structure. C-band data was largely influenced by foliage volume and therefore may be useful for monitoring defoliation. Changes in water content modulated the C-band signal by <1 dB which may be insufficient for a meaningful retrieval of drought effects on forest. C-band sensitivity to significant clear-cuts was rather low (1.5 dB). More subtle effects such as selective logging or thinning may not be easily detected using C- or L-band data with the longer P-band needed for retrieving small intensity forest disturbances. Overall, the simulations emphasize that additional effort is needed to overcome current limitations arising from the use of a single frequency, acquisition time and geometry by tapping the advantages of dense time series, and by combining acquisitions from active and passive sensors. The simulation results may be applicable to forests outside of Romania since the forests types used in the study have similar morphological characteristics to forests elsewhere in Europe.

Ozone risk assessment is affected by nutrient availability: Evidence from a simulation experiment under free air controlled exposure (FACE).

Assessing ozone (O3) risk to vegetation is crucial for informing policy making. Soil nitrogen (N) and phosphorus (P) availability could change stomatal conductance which is the main driver of O3 uptake into a leaf. In addition, the availability of N and P could influence photosynthesis and growth. We thus postulated that the sensitivity of plants to O3 may be changed by the levels of N and P in the soil. In this study, a sensitive poplar clone (Oxford) was subject to two N levels (N0, 0 kg N ha-1; N80, 80 kg N ha-1), three P levels (P0, 0 kg P ha-1; P40, 40 kg P ha-1; P80, 80 kg P ha-1) and three levels of O3 exposure (ambient concentration, AA; 1.5 × AA; 2.0 × AA) for a whole growing season in an O3 free air controlled exposure (FACE) facility. Flux-based (POD0 to 6) and exposure-based (W126 and AOT40) dose-response relationships were fitted and critical levels (CLs) were estimated for a 5% decrease of total annual biomass. It was found that N and P availability modified the dose-response relationships of biomass responses to O3. Overall, the N supply decreased the O3 CLs i.e. increased the sensitivity of poplar to O3. Phosphorus alleviated the O3-caused biomass loss and increased the CL. However, such mitigation effects of P were found only in low N and not in high N conditions. In each nutritional treatment, similar performance was found between flux-based and exposure-based indices. However, the flux-based approach was superior, as compared to exposure indices, to explain the biomass reduction when all nutritional treatments were pooled together. The best O3 metric for risk assessments was POD4, with 4.6 mmol m-2 POD4 as a suitable CL for Oxford poplars grown under various soil N and P conditions.

Unique postglacial evolution of the hornbeam (Carpinus betulus L.) in the Carpathians and the Balkan Peninsula revealed by chloroplast DNA.

The Balkan Peninsula is one of the largest and most important European glacial refugia. However, the evolutionary history and phylogeographic pattern of temperate tree species that survived in the Balkans glacial refugia and their contribution to the genetic structure of the current population in the Carpathian Mountains remains poorly understood. Using polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP), and extensive population sampling, we explored the phylogeographic pattern of Carpinus betulus in both the Balkan Peninsula and the Carpathian region. We aimed to determine the locations of potential glacial refugia, in order to delineate post-glacial colonization routes, and also to test if northern cryptic refugia had persisted during the Last Glacial Maximum (LGM). Our results provide strong support for the existence of multiple refugia similar to the 'refugia-within-refugia' scenario, which would suggest that Carpinus betulus has experienced a complex evolutionary history. In agreement with pollen data, our findings corroborate with previous hypotheses suggesting that hornbeam has a distinct postglacial evolution in the southeast of Europe (Carpathians and Balkan Peninsula) compared with that in Western Europe. Three postglacial re-colonization routes, from three distinct effective glacial refugia: (1) the Dinaric Alps; (2) the Pirin and Rhodope Mountains and (3) the Strandzha Mountains, were detected within the Balkan Peninsula. The pattern of cpDNA haplotypes distribution across the Ukrainian Carpathians revealed a "suture zone", which is a consequence of contact due to postglacial re-colonization between hornbeam populations originating from all three effective glacial refugia. The peculiar haplotype geographical structure found in C. betulus and the identified multiple glacial refugia in the Balkans may have direct consequences in the management and conservation of hornbeam forest genetic resources.

A new-generation 3D ozone FACE (Free Air Controlled Exposure).

To artificially simulate the impacts of ground-level ozone (O3) on vegetation, ozone FACE (Free Air Controlled Exposure) systems are increasingly recommended. We describe here a new-generation, three-dimensional ozone FACE, with O3 diffusion through laser-generated micro-holes, pre-mixing of air and O3, O3 generator with integral oxygen generator, continuous (day/night) exposure and full replication. Based on three O3 levels and assumptions on the pre-industrial O3 levels, we describe principles to calculate relative yield/biomass and estimate impacts even at lower-than-ambient O3 levels. The case study is called FO3X, and is at present the only ozone FACE in Mediterranean climate and one of the very few ozone FACEs investigating more than one stressor at a time. The results presented here will give further impulse to the research on O3 impacts on vegetation all over the world.